Anticonvulsant activity of reaction products of 5,5-diphenylhydantoin with substituted methylene bromides

Article abstract of DOI:10.1002/1521-4184(200112)334:11<366::AID-ARDP366>3.0.CO;2-Q

Some derivatives of 5,5-diphenylhydantoin (phenytoin) were synthesized by the alkylation of phenytoin with substituted methylene bromides. The hydantoins were evaluated for possible anticonvulsant activity in the maximal electroshock (MES)- and subcutaneous pentylenetetrazole (ScMet)-induced seizures and for neurotoxicity in the rotorod test in mice and rats.

Full text of DOI:10.1002/1521-4184(200112)334:11<366::AID-ARDP366>3.0.CO;2-Q

Notes
Cyril O. Usifoh
Anticonvulsant activity of reaction products
of 5,5-diphenylhydantoin with substituted
methylene bromides
Department of Pharmaceutical
Chemistry, Faculty of Pharmacy,
University of Benin, Nigeria
Some derivatives of 5,5-diphenylhydantoin (phenytoin) were synthesized by the alkyla-
tion of phenytoin with substituted methylene bromides. The hydantoins were evaluated
for possible anticonvulsant activity in the maximal electroshock (MES)- and subcutane-
ous pentylenetetrazole (ScMet)-induced seizures and for neurotoxicity in the rotorod test
in mice and rats.
Key Words: Anticonvulsants; Hydantoins; Methylene bromides
Received: July 4, 2001 [FP608]
Introduction
Epilepsy is a symptom of excessive temporary neuronal
discharge, characterized by discrete recurrent episodes, in
which there is a disturbance of movement, sensation,
behavior, perception, and/or consciousness. It has been
estimated that 0.5 to 1% of the population is affected by
some form of epilepsy [1,2]
.
Anticonvulsant drugs display their activity by different
mechanisms. The drugs can influence the ion transport
across cell membranes, or inhibitory or excitatory neuro-
transmitter systems [3]
.
Although many treatments for epilepsy are available, more
efforts should be devoted to novel approaches. The quest[4]
to discover more efficacious new anticonvulsant drugs with
fewer adverse effects and the potential to provide the
patient with a better quality of life prompted the preparation
and biological evaluation of the derivatives of hydantoins.
The compounds obtained were evaluated for anticonvul-
sant activity in the maximal electroshock (MES)- and sub-
cutaneous pentylenetetrazole (ScMet)-induced seizures
and for neurotoxicity in the rotorod test in mice and rats.
Scheme. i = K2CO3, DMF, 25 °C; or KOH, EtOH, reflux;
ii = MeOH, reflux, 5 h.
Chemistry
The hydantoins were synthesized as outlined in the scheme
below. Alkylation of N-3 of the hydantoin was accomplished
with the appropriate methylene bromide. In the synthesis
of 5,5-diphenyl-3-(acetonitrilyl)-hydantoin 3b, the yield was
higher when alkylation was executed with potassium hy-
droxide. The removal of the trimethyl moiety from 3c to give
4 was accomplished in refluxing methanol. This is in agree-
ment with a recent study on the de-protection or removal
of the triphenylmethyl moiety[5]. The assignment of all
structures was established on the basis of IR, NMR, mass
spectrometry, and microanalysis.
Table 1. Physical data.
————————————————————————————–
Cmpd Reaction
time
Crystallizing
solvent
Melting
point
————————————————————————————–
3a
4 h
Ethanol
137–138 °C
(Lit[6]:137–138°C)
3b
3c
4
36 h
30 h
5 h
Hexane–Ethyl acetate 194–195 °C
Hexane–Ethyl acetate 180–181 °C
Hexane–Ethyl acetate 237–239 °C
————————————————————————————–
Pharmacology
———
Correspondence: Dr. Cyril O. Usifoh, Department of Pharmaceu-
tical Chemistry, Faculty of Pharmacy, University of Benin, Nigeria.
E-mail: usifoh@uniben.edu.ng
The four compounds (3a–c, 4) were screened for anticon-
vulsant activity according to standard procedures, which
involves the maximal electroshock seizure test (MES test)
Fax: +234 52 602257
© WILEY-VCH Verlag GmbH, D-69451 Weinheim, 2001
0365-6233/01/1111/0366 $17.50 +.50/0

Arch. Pharm. Pharm. Med. Chem. 2001, 334, 366–368
Phenytoin derivatives 367
Table 2. Anticonvulsant activity in the MES test and the ScMet test and toxicity in the rotorod test of
hydantoins following intraperitoneal administration to mice. The numbers are expressed as animals
protected/animals tested.
——————————————————————————————————————————————
Cmpd
Dose
[mg/kg]
MES
0.5 h
ScMet
Toxicity
0.5 h 4 h
Class a)
4 h
0.5 h
4 h
——————————————————————————————————————————————
3a
30
100
300
3
10
30
100
300
30
100
300
30
0/1
0/3
0/1
–
0/1
3/3
1/1
0/4
0/4
1/1
3/3
1/1
0/1
0/3
1/1
0/1
0/3
0/1
0/1
0/1
0/1
–
0/1
0/1*
0/1*
–
0/4
0/8
0/4
–
0/2
0/4
2/2
0/4
0/4
0/2
2/4
1/2
0/2
0/4
0/2
0/2
0/4
0/2
1
3b
1
–
–
–
–
0/1
1/3
1/1
0/1
0/3
0/1
0/1
0/3
0/1
0/1
0/1
0/1
0/1
0/1*
0/1*
0/1
0/1
0/1
0/1
0/1
0/1
0/1*
0/1*
0/1
1/5*
1/5*
0/1
0/4
0/8
2/4
0/4
0/8
0/4
0/4
0/8
0/4
3c
4
3
1
100
300
——————————————————————————————————————————————
*Death following clonic seizures. a) Classification according to reference [7]. Class 1, active at 100 mg/kg;
Class 2, active at 300 mg/kg; Class 3, inactive.
Additional evaluation of 3a and 3b in rats at a dose of
30 mg/kg (p.o.) at MES over a period of 4 h revealed the
time for peak activity. Both compounds were significantly
not neurotoxic in the rotorod test. The time for peak activity
for 3a was 4 h while 3b displayed more or less the same
level of activity throughout the period. No anticonvulsant
activity was however observed for 3b after 50 min.
Table 3. Quantitative anticonvulsant (TD50, ED50 MES, and PI)
activity of 3b and standard anticonvulsants after intraperitoneal
administration to mice. The values are expressed in mg/kg. 95%
confidence intervals are listed in brackets, while the time of test is
listed in square brackets.
————————————————————————————–
Cmpd
TD50
ED50
PI
(rotorod)
(MES)
(TD50/
ED50
)
————————————————————————————–
However, in comparison to some known antiepileptic drugs,
the protective index (PI) in the MES test for 3b and val-
proate is the same. The median toxic dose (TD50) for 3b as
well as effective dose (ED50) were correspondingly high
compared to phenytoin. The median toxic dose (TD50) of
3b was 2-fold of phenytoin while the median effective dose
was 7.5-fold less active than phenytoin. The modification
of phenytoin by alkylation with subtituted methylene bro-
mides therefore did not appreciably enhance the activity of
phenytoin.
3b
82.4 (68.8–98.6)
[2h]
48.8 (34.6–62.7)
[2h]
6.48 (5.65–7.24)
[2h]
287 (237–359)
[1/4h]
1.7
6.6
1.7
Phenytoin a) 42.8 (36.4–47.5)
[2h]
Valporate a) 483 (412–571)
[1/4h]
————————————————————————————–
a) Data from references [7,9].
Table 4. Anticonvulsant activity of 3a and 3b. Test results in rats
(30 mg/kg p.o.).
————————————————————————————–
Cmpd Test
Time (h)
0.50 1.0
————————————————————————————–
Acknowledgments
0.25
2.0
4.0
The anticonvulsant and toxicity testing by Dr. J. P. Stables at
the NINDS Epilepsy Branch, National Institutes of Health,
Rockville, MD, USA is gratefully acknowledged.
3a
MES
TOX
0/4
0/4
0/4
1/4
0/4
0/4
0/4
0/4
4/4
0/4
3b
MES
TOX
2/4
0/4
0/4
0/4
1/4
0/4
2/4
0/4
2/4
0/4
————————————————————————————–
Experimental
and the seizure threshold test with subcutaneous pentyle-
netetrazole (scMet test). The acute neurological toxicity
was determined in the rotorod test. The results of the
screening in mice are summarized in Table 2. Only com-
pounds 3a and 3b displayed an effect in the maximal
electroshock seizure test at a dose of 100 mg/kg with 3b
also exhibiting an effect at a lower dose of 30 mg/kg. Fifty
percent of the numbers of mice used were neurotoxic at a
minimum dose of 100 mg/kg. Death following clonic sei-
zures was recorded in some cases with the subcutaneous
pentylenetetrazole (scMet test). The anticonvulsant activity
recorded for compound 4 in the scMet test with correspond-
ing absence of acute neurotoxicity was low.
Chemistry
Melting points were determined with a Kofler hot-stage microscope
and were reported uncorrected. The reactions and purity of the
products were monitored by TLC. Nuclear Magnetic Resonance
spectra were recorded on a Varian Gemini 200 (TMS); infra-red
spectra were measured on a Perkin-Elmer type 457 and the mass
spectra were determined using a Varian MAT 44S, EI: 70 eV.
Propargyl bromide and bromoacetonitrile were obtained from
Aldrich while N-(triphenyl)-5-[4-(bromomethyl)-biphenyl-2-yl]-
tetrazole was synthesized according to a known procedure[8]
.

368 Phenytoin derivatives
Arch. Pharm. Pharm. Med. Chem. 2001, 334, 366–368
51(9). C29H22N6O2 (486.54). Calc. C 71.59 H 4.56 N 17.27;
found C 71.50 H 4.44 N 17.14.
Alkylation of 5,5-diphenyl-3-prop-2-ynylhydantoin
Method I: A mixture of substituted methylene bromide
(0.02 mol), 5,5-diphenyl-3-prop-2-ynylhydantoin (0.02 mol),
and potassium hydroxide (0.02 mol) was refluxed in 30 ml
ethanol. Standard work-up procedures was carried out and the
compound crystallized from appropriate solvent.
Pharmacology
Anticonvulsant testing was provided by the Antiepileptic Drug
Development Program, Epilepsy Branch, Division of Convul-
sive, Developmental and Neuromuscular Disorders, National
Method II: Substituted methylene bromide (3.97 mmol) was
added to a stirred suspension of 5,5-diphenyl-3-prop-2-ynylhy-
dantoin (3.97 mmol) and potassium carbonate (3.97 mmol) in
20 ml N,N-dimethylformamide at room temperature. Standard
work-up procedures were executed and the compound crystal-
lized from appropriate solvent.
[6]
Institutes of Health, according to standard procedures
and
included the MES test and the seizure scMet test. In the MES
test, an electrical stimulus of 50 mA was delivered for 0.2 sec
via corneal electrodes to mal CF1 mice at 30 minutes and 4 h
after the administration of the compounds. Blockade of the tonic
extension of the hind limbs was considered protection against
seizures. For the scMet test a convulsant dose of 85 mg/kg of
pentylenetetrazole dissolved in saline was injected in a loose
fold of skin on the back of the neck and the animals were
isolated and observed for 30 minutes. Absence of clonic
spasms for at least 5 sec indicated the elevation of the penty-
lenetetrazole-induced seizure threshold. The acute neurologi-
cal toxicity was determined in the rotorod test where the animal
was placed on a rod rotating at 6 rpm. Neurological deficiency
was indicated by inability to maintain equilibrium for 1 min in
each of 3 trials. For all these evaluations the compounds were
dissolved or suspended in 0.5 % aqueous methyl cellulose.
5,5-Diphenyl-3-prop-2-ynylhydantoin 3a
Method I: Propargyl bromide was added to 5,5-diphenyl-3-prop-
2-ynylhydantoin and gave after work-up fine needles of 5,5-
diphenyl-3-prop-2-ynylhydantoin 3a (5.1 g, 76%).
5,5-Diphenyl-3-(acetonitrilyl)-hydantoin 3b
Method II: 0.92 g (80%) ; IR (KBr): ν (cm–1) = 3390 (NH), 1775,
1730 (C=O), 1600; 1H NMR (Acetone d6): δ = 4.72 (s, 2H, CH2),
7.48 (brs, 10H, Ar-H), 8.89 (brs, 1H, N-H). MS: m/z = 292 [M++
1] (12), 291 [M+] (60), 262 (69), 235 (7), 208 (20), 180 (100),
165 (26), 152 (6), 104 (21), 77 (21), 51 (9); C17H13N3O2 (291.31
). Calc. C 70.09 H 4.50 N 14.43; found C 70.20 H 4.54 N 14.30.
References
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1
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7.57 (t, J = 7.8Hz, 2H, Ar-H), 7.67 (d, J = 7.8Hz, 2H, Ar-H), 9.80
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